Adenosine is known to be associated with effects such as inhibition of immune response, coronary vasodilation, stimulation of angiogenesis, and inhibition of inflammatory reactions. Some authors suggest that adenosine may also have similar functions in tumor tissues. Tissue levels of adenosine are under close regulation by different enzymes acting at different levels. Adenosine is produced from AMP by the action of 5'-nucleotidase (5'-NT) and is converted back into AMP by adenosine kinase (AK) or into inosine by adenosine deaminase (ADA). Inosine is converted into purine catabolites by purine nucleoside phosphorylase (PNP), whereas AMP is converted into ADP and ATP by adenylate kinase (MK). The aim of this study was to analyze the activities of the above enzymes in fragments of neoplastic and apparently normal mucosa, obtained less than 5 cm and at least 10 cm from tumors, in 40 patients with colorectal cancer. The results showed much higher activities of ADA, AK, 5'-NT, and PNP in tumor tissue than in neighboring mucosa (p > 0.01 for ADA, AK, and PNP; p > 0.05 for 5'-NT), suggesting that the activities of purine metabolizing enzymes increase to cope with accelerated purine metabolism in cancerous tissue. The simultaneous increase in ADA and 5'-NT activities might be a physiological attempt by cancer cells to provide more substrate to accelerate salvage pathway activity.
The metabolism of purine nucleotides was studied in human peripheral blood lymphocytes from healthy subjects and patients with B-cell chronic lymphocytic leukemia. Nucleotide content was determined by HPLC. The rate of de novo synthesis of purine nucleotides was measured kinetically by following the incorporation of 14C-formate into the nucleotides of a lymphocyte suspension. The patterns of the main enzymes involved in purine nucleotide metabolism (those of the salvage pathway and catabolism) were estimated by a radiochemical method. Although the data expressed in relation to cells and protein showed some discrepancies, several common differences were evident in both cases. The main differences were an increase in NAD and IMP, a sharp decrease in 5'-nucleotidase activities and in total guanylate content and synthesis, and an increase in the A/G ratio in lymphocytes of patients with respect to controls. The changes in these parameters in CLL indicate an imbalance in purine metabolism and may play a specific role in the biology of the leukemia cell. They are also potential biochemical markers of lymphoid malignancies and may be useful in chemotherapic applications.
Background:The diagnosis and monitoring of severe combined immunodeficiency disease (SCID) attributable to adenosine deaminase (ADA) deficiency requires measurements of ADA, purine nucleoside phosphorylase (PNP), and S-adenosyl-L-homocysteine-hydrolase (SAHH) activity and of deoxyadenosine metabolites. We developed capillary electrophoresis (CE) methods for the detection of key diagnostic metabolites and evaluation of enzyme activities. Methods: Deoxyadenosine metabolites were separated in 30 mmol/L sodium borate-10 mmol/L sodium dodecyl sulfate (pH 9.80) at 25°C on a 60-cm uncoated capillary. For determination of enzyme activities, substrate-product separation and measurements were carried out in 20 mmol/L sodium borate (pH 10.00) at 25°C on a 42-cm uncoated capillary. Results: Deoxynucleotides and deoxyadenosine were readily detectable in erythrocytes and urine, respectively. Both methods were linear in the range 2-500 mol/L (r >0.99). Intra-and interassay CV were <4%. Enzyme activities were linear with respect to sample amounts in the incubation mixture and to incubation time (r >0.99 for both). In erythrocytes from healthy individuals, mean (SD) ADA activity was 5619 (2584) nmol/s per liter of packed cells. In erythrocytes of SCID patients at diagnosis, ADA activity was 56.9 (48.3) nmol/s per liter of packed cells; SAHH activity was also much reduced. PNP activity was similar in patients and controls. Conclusions: CE can be used to test ADA deficiency and enables rapid assessment of ADA expression in hema-
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